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Gene/Clone	Species	Stage	Anatomy Item	Experimenter
pax6	xenopus

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Experiment details for pax6

Rodríguez-Seguel E et al. (2009) Assay

The Xenopus Irx genes are essential for neural patterning and define the border between prethalamus and thalamus through mutual antagonism with the anterior repressors Fezf and Arx.

Gene	Clone	Species	Stages	Anatomy
pax6.L		laevis	NF stage 14 to NF stage 15	neural plate , anterior neural fold , anterior neural ridge

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Fig. 1. Expression pattern of Xenopus laevis Irx genes. (A1–5) Vegetal views at early gastrula show that Irx1 (A1) and Irx3 (A3) are expressed in two dorso-lateral mesodermal bands (arrowhead). In addition, Irx1 and Irx2 are expressed in the presumptive neural ectoderm (A1 and A2, insets). (B1–5) Dorsal views of early neurula embryos illustrating the similar expression domains of all Irx genes (n, notochord). Arrowheads in (B1) and (B2) point at the placodal expression of Irx1 and Irx2. (C1) Early neurula embryo double-stained for Six3 (purple) and Irx1 (cyan) genes. The expression patterns of these genes are separated a few cell diameters (arrowheads). (C2–C5) Early neurula embryo double-stained for Irx genes (purple) and Fezf2 (cyan). Inset in (C2) show a similar embryo double-stained for Irx1 (purple) and Fezf2 (cyan). Insets in (C3–5) show double in situs developed in a single colour (purple) for Fezf2 and Irx3 (C3), Irx4 (C4) and Irx4 (C5). All Irx genes show similar anterior border abutting Fezf expression (arrowheads). (D1, D2) Double staining for Irx1 and Irx2 (D1) or Irx3 (D2) also show that these genes share their anterior expression limit (arrowheads). (D3, D4). This border is rostral to Pax2, as shown in double-stained embryos for this gene and Irx1 (D3) or Irx3 (D4). Inset in (D3) show a sagittal section of a double-labelled embryo for Pax2 (purple) and Irx1 (cyan). Note that Irx1 is anterior to that of Pax2. (D5) The Irx anterior limit overlaps with the posterior expression of Pax6 in the forebrain. (E1–G5) At tailbud stage, in lateral views (E1–5) or in dorsal views (F1–5, G1–5) all Irx genes show similar expression patterns in the brain although with different intensities in different regions. This is more clearly seen in double-stained embryos for Irx genes (purple) and Krox20 (cyan) (G1–G5). Red, black and blue arrowheads point at the M/H boundary, rhombomere 3 (r3) and rhombomere 5, respectively. Green arrowheads and red arrows point at the otic vesicle and pronephros, respectively. Stg, stage.

Gene	Clone	Species	Stages	Anatomy
pax6.L		laevis	NF stage 16	diencephalon , hindbrain , pre-chordal neural plate , chordal neural plate , anterior neural fold

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Fig. 3. Irx genes are required for neural patterning. All panels, except (M) that show a stage 18 embryo, show dorsal views of stage 14–16 embryos co-injected with a mix of all five Irx MOs and LacZ mRNA. Black and red arrowheads point at the control or injected side, respectively. (A–C) Forebrain markers Rx1, Six2 and Fezf2 are expanded posteriorly (compare red with black arrowheads). (D) In addition, the anterior Pax6 domain is also shifted caudally at the expenses of the midbrain territory that lack the expression of this gene (compare red with black arrowhead). (E–F) The expression of Wnt1 and Irx3 in the posterior diencephalon is strongly downregulated (compare red with black arrowheads). (G–I) The midbrain is also reduced in the Irx morphant, as determined by the expression of En2, Pax2 and Wnt4. In addition, Wnt4 expression in the spinal cord in strongly impaired (I). (J–L) The rhomboencephalon markers Gbx2, Krox20 and Nhf1β are also downregulated in the injected embryos. (M) At stage 18, Sox2 expression shows and altered morphology of brain structures in the Irx impaired side. (N, O) The proneural gene Ngnr1 (N) and the primary neurons markers Ntubulin (O) are also downregulated in the injected side.